A strain of Lactobacillus paracasei and its application in improving excessive facial oiliness

Abstract

The application belongs to the technical field of microorganism screening and application, and particularly relates to a lactobacillus paracasei and application thereof in improving the problem of excessive facial oil secretion. The lactobacillus paracasei has a preservation number of CCTCC NO: M2021587, strong acid resistance and strong antioxidant capacity, can effectively relieve skin damage caused by various factors, reduce the problem of excessive facial skin oil secretion, and can be widely applied to skin care products.

Description

Technical Field

[0001] The invention relates to the technical field of functional microorganism screening and application, and in particular to a Lactobacillus paracasei and application thereof in improving the problem of excessive oil production on the face. Background Art

[0002] Human skin is home to trillions of microorganisms, including bacteria, fungi, and protozoa. These microorganisms, along with surface cells, tissues, their secretions, and the skin microenvironment, form an ecosystem. Like the intestinal flora, the skin microbiome plays a crucial role in skin health, helping the body fight foreign pathogens, breaking down skin cell metabolites, and regulating the immune system. Common skin health conditions, such as acne, psoriasis, and atopic dermatitis, are closely linked to an imbalance in the skin's microbiome. In recent years, skin microbiota has become a focus of attention in the cosmetics and academic communities. Probiotic fermentation lysate, a cosmetic ingredient produced through fermentation and homogenization of a bacterial strain, includes bacterial components such as exopolysaccharides, peptidoglycan, cell membrane proteins, and lipoteichoic acid, as well as metabolites secreted by probiotic cells, such as lactic acid, acetic acid, intracellular polysaccharides, lipids, bacteriocins, and various small peptides. For example, studies have found that the fermentation filtrate of lactobacillus with mung bean seed extract and monosodium glutamate can utilize the dual advantages of mung bean and monosodium glutamate, and has soothing, anti-aging and moisturizing effects, which can enhance the skin barrier function; after the Chinese medicinal material Dendrobium officinale is fermented by lactic acid bacteria, its protein and fat content is reduced, its antioxidant and anti-aging effects are improved, its skin absorption and utilization are improved, and its anti-aging and moisturizing effects are enhanced; Silva et al. found that the fermentation lysate of bifidobacterium can inhibit common skin pathogens such as Staphylococcus aureus, Staphylococcus epidermidis, Candida albicans and Malassezia furfur; in addition, the fermentation lysate of bifidobacterium also has anti-inflammatory effects, which can inhibit the release of pro-inflammatory cytokines such as IL-6, IL-1β and TNF-α by keratinocytes and skin fibroblasts.

[0003] Skin lipids are one of the important components of the skin and play a vital role in maintaining the skin barrier function, preventing transepidermal water loss and the invasion of external environmental substances. When the composition or content of skin lipids changes, the normal physiological function of the skin, especially the skin barrier function, will be affected. The lipids on the surface of the skin can be divided into sebaceous gland lipids and intercellular lipids according to their sources. Sebaceous gland lipids are directly secreted by the sebaceous glands to the surface of the skin, while intercellular lipids are mainly dispersed between the stratum corneum cells as the cells grow and metabolize, and finally distributed to the skin surface as the keratinocytes disintegrate. The sebum secretion rate of a normal adult is 1mg / 10cm 2When the value of / 3h exceeds the value, oily skin appearance is prone to occur, and people with excessive sebum secretion are prone to have skin health problems such as pore blockage, comedones, blackheads and acne. In addition, oily skin has an unbalanced water content and a low pH value, and the skin is prone to have an oily appearance, and people with oily skin are prone to have a frustrated and embarrassed psychology. Studies have shown that oily skin can have a significant negative impact on the quality of life of Chinese women, and oil control cosmetics have always been a hot spot for consumers.

[0004] The oil control is interpreted in the Cosmetics Classification Rules and Classification Catalogue as "helping to slow down the sebum secretion and deposition of the application site, or making the sebum secretion of the application site not obvious". Products for removing excess sebum on the skin are generally mainly of the cleaning type and the adsorption type. However, some cleaning products can make the skin excessively dry, and the sebum can be excessively compensated, thereby aggravating the problem of excessive oil on the skin surface. In addition, medical and aesthetic methods such as chemical peeling by glycolic acid substances or injection of botulinum toxin to reduce sebum secretion can also alleviate the problem of excessive oil on the skin. However, the high price of medical and aesthetic products and the inconvenience of daily care have affected consumers' facial oil control care. Therefore, it is of great social and economic benefit to develop a strain of lactic acid bacteria that can improve the problem of excessive oil on the face. SUMMARY

[0005] The purpose of the present application is to provide a strain of Lacticaseibacillus paracasei and its application in improving the problem of excessive oil on the face. The Lacticaseibacillus paracasei has strong acid tolerance and strong antioxidant capacity, can effectively alleviate skin damage caused by various factors, and can alleviate the problem of excessive oil on the face, and can be widely used in skin care products.

[0006] In one aspect, the present application relates to a strain of Lacticaseibacillus paracasei, which is named Lacticaseibacillus paracasei VHProbi E12 (Lacticaseibacillus paracasei VHProbi E12), and was preserved in the China Center for Type Culture Collection of Wuhan University in Wuhan, China on May 24, 2021, and the preservation number is CCTCC NO: M2021587.

[0007] In one aspect, the present application relates to the application of the fermentation lysate of the Lacticaseibacillus paracasei VHProbi E12 in improving the problem of excessive oil on the face.

[0008] The preparation method of the fermentation lysate of the Lacticaseibacillus paracasei VHProbi E12 comprises the following steps:

[0009] (1) Preparation of fermentation broth

[0010] Under sterile conditions, Lactobacillus paracasei VHProbi E12 seed liquid was inoculated into a fermentation medium at a volume ratio of 3%, and cultured at 37° C. for 24 hours at a rotation speed of 100 rpm, a ventilation volume of 1 L / min, a tank pressure of 0.05 MPa, and a fermentation endpoint pH of 3.5-4.2 to prepare a Lactobacillus paracasei VHProbi E12 fermentation liquid;

[0011] (2) Emulsification and homogenization of fermentation broth

[0012] The fermentation broth of Lactobacillus paracasei VHProbi E12 was emulsified and homogenized using a homogenizer at a pressure of 1000 bar for 3 times, and then placed in an 80°C water bath for 30 minutes to completely kill the living cells in the fermentation broth, thereby obtaining a fermentation lysate of Lactobacillus paracasei VHProbi E12.

[0013] The components and contents of the fermentation medium are: 25 g / L sucrose, 30 g / L yeast powder, 20 g / L soy peptone, 0.10 g / L magnesium sulfate, and 0.05 g / L manganese sulfate.

[0014] The present invention also relates to application of the Lactobacillus paracasei VHProbi E12 fermentation lysate in skin care products.

[0015] The present invention also provides a skin care product comprising a fermentation lysate of Lactobacillus paracasei VHProbi E12.

[0016] The skin care product further comprises any one or more of butylene glycol, mineral oil, glyceryl caprate, cetyl alcohol, dimethicone, peg-100 stearate, 1,2-hexanediol, hydroxyacetophenone, phenoxyethanol, glyceryl stearate, carbomer, arginine, and disodium edta.

[0017] The components and their mass percentages in the skin care product are: 8% of Lactobacillus paracasei VHProbi E12 fermentation lysate, 5% of butylene glycol, 2% of mineral oil, 1.5% of capric glyceryl, 1.5% of cetyl alcohol, 0.5% of dimethicone, 0.5% of PEG-100 stearate, 0.4% of 1,2-hexanediol, 0.3% of hydroxyacetophenone, 0.25% of phenoxyethanol, 0.2% of glyceryl stearate, 0.15% of carbomer, 0.13% of arginine, 0.05% of disodium edta and 79.52% of water.

[0018] The present invention also provides application of the skin care product in improving the problem of excessive oil production on facial skin.

[0019] The Lactobacillus paracasei VHProbi E12 fermentation lysate provided by the present invention can inhibit 5α-reductase activity, reduce skin lipid secretion, and alleviate oily skin.

[0020] The Lactobacillus paracasei VHProbi E12 fermentation lysate provided by the present invention has a significant inhibitory effect on the proliferation of SZ95 human sebaceous gland cells, and can reduce the problem of excessive sebum secretion caused by excessive proliferation of SZ95 human sebaceous gland cells; at the same time, the Lactobacillus paracasei VHProbi E12 fermentation lysate can activate TRPV1 and AMPK signaling pathways, inhibit the excessive synthesis of lipids in SZ95 human sebaceous gland cells induced by linoleic acid, thereby effectively reducing the oil secretion of SZ95 human sebaceous gland cells and improving the problem of excessive oil secretion of facial skin.

[0021] Human efficacy trials confirmed that after 60 days of using a moisturizer containing Lactobacillus paracasei VHProbi E12 fermentation lysate, skin oil content decreased by 15.30%, porphyrin levels decreased by 30.74%, and skin oiliness was alleviated. Changes in facial porphyrin levels, measured using a VISIA-CR instrument, showed a significant decrease compared to baseline, indicating less oily skin. 80.4% of participants reported a reduction in facial oiliness, demonstrating a significant improvement. BRIEF DESCRIPTION OF THE DRAWINGS

[0022] Figure 1 This is the result of the SZ95 human sebaceous gland cell activity inhibition test;

[0023] Figure 2 This is a comparison chart of lipid synthesis rates;

[0024] Figure 3 is a comparison of TRPV1 and AMPK mRNA expression levels;

[0025] Figure 4 This is a comparison chart of the oil content in the stratum corneum of the subjects;

[0026] Figure 5 This is a graph showing changes in rhodopsin content on the subjects' faces. DETAILED DESCRIPTION

[0027] The screening methods of the present invention are not limited to those described in the Examples; any known method that can achieve the screening purpose may be used. The screening descriptions in the Examples are merely illustrative of the present invention and are not intended to limit the scope of protection of the present invention. Modifications or substitutions made to the methods, steps, or conditions of the present invention without departing from the spirit and substance of the present invention are within the scope of the present invention.

[0028] The Lactobacillus paracasei VHProbi E12 described in the embodiments of the present invention was isolated from the feces of a healthy infant and was deposited in the China Center for Type Culture Collection of Wuhan University, Wuhan, China on May 24, 2021, with the deposit number being CCTCC NO: M2021587.

[0029] Lactobacillus paracasei VHProbi E12 has strong acid resistance; does not produce hemolysin, does not dissolve blood cells, is sensitive to common antibiotics such as erythromycin and tetracycline, and has good biosafety; can tolerate higher salinity, with a maximum tolerable salt concentration of 5%.

[0030] Lactobacillus paracasei VHProbi E12 has strong antioxidant capacity and can effectively scavenge DPPH free radicals with a scavenging rate of 34.44%, which is significantly higher than that of the control strain Lactobacillus paracasei IMC-4.

[0031] Lactobacillus paracasei VHProbi E12 can effectively promote skin cell proliferation.

[0032] Lactobacillus paracasei VHProbi E12 significantly reduces cell damage caused by Staphylococcus aureus infection, providing strong protection. While the cell mortality rate in the control group infected with S. aureus reached 55.7%, the mortality rate in the probiotic-treated group, which also included heat-killed Lactobacillus paracasei VHProbi E12, dropped to 37.0% (P<0.005), demonstrating a significant effect.

[0033] Lactobacillus paracasei VHProbi E12 effectively reduces skin cell damage caused by UVB radiation. Compared with the control group, the viability of cells in the UVB-damaged group decreased significantly, to only 49.1%, while the viability of cells in the probiotic-treated group, which had been pretreated with inactivated Lactobacillus paracasei VHProbi E12, reached a high of 72.6% (P < 0.05).

[0034] Lactobacillus paracasei VHProbi E12 effectively reduces cell damage caused by hydrogen peroxide oxidation. Compared with the control group, the viability of cells in the hydrogen peroxide-damaged group dropped significantly to 49.8%. However, the viability of cells in the probiotic-treated group, which had been pretreated with inactivated Lactobacillus paracasei VHProbi E12 bacteria, reached 59.4%, a significant improvement (P < 0.005).

[0035] Lactobacillus paracasei VHProbi E12 can effectively alleviate skin damage caused by hydrogen peroxide oxidation, exhibiting significant anti-inflammatory and immunomodulatory effects. Compared with the control group, skin cell viability was significantly reduced after hydrogen peroxide application, to only 50.6%, while skin cell secretion of three pro-inflammatory cytokines, IL-1α, IL-1β, and IL-8, was significantly increased. In the probiotic-treated group, which was pretreated with inactivated Lactobacillus paracasei VHProbi E12 bacteria before hydrogen peroxide application, skin cell viability reached 57.2% (P < 0.05), and secretion of IL-1α, IL-1β, and IL-8 was also significantly reduced (P < 0.005), demonstrating a highly significant effect.

[0036] Lactobacillus paracasei VHProbi E12 effectively reduces the damage caused by Triton X-100 to the skin. Compared to the control group, the transmembrane electrical resistance of a skin model treated with a 0.1% Triton X-100 solution decreased by 61.4%. However, the decrease in transmembrane electrical resistance in the probiotic-treated group, which had been pretreated with inactivated Lactobacillus paracasei VHProbi E12 bacteria, was less pronounced, decreasing by only 52.4% compared to the control group (P < 0.05).

[0037] Lactobacillus paracasei VHProbi E12 has a certain protective effect against skin damage caused by various factors such as bacterial infection, ultraviolet rays, oxidative damage and chemical damage. It can be widely used in cosmetics or medicines and has broad application prospects.

[0038] The present invention will be further described below with reference to specific embodiments.

[0039] Example 1 Preparation of Lactobacillus paracasei VHProbi E12 fermentation lysate

[0040] 1.1 Preparation of fermentation broth

[0041] Under sterile conditions, Lactobacillus paracasei VHProbi E12 seed liquid is inoculated into a fermentation medium at a volume ratio of 3%, wherein the components and their contents in the fermentation medium are respectively: 25 g / L sucrose, 30 g / L yeast powder, 20 g / L soy peptone, 0.10 g / L magnesium sulfate and 0.05 g / L manganese sulfate, and the culture is carried out at 37°C for 24 hours, with a rotation speed of 100 rpm, a ventilation volume of 1 L / min, a tank pressure of 0.05 MPa, and a fermentation endpoint pH of 3.5-4.2 to prepare a fermentation liquid;

[0042] 1.2 Emulsification and homogenization of fermentation broth

[0043] The fermentation broth was emulsified and homogenized using a SPXFLOW homogenizer (model: APV-1000) at a pressure of 1000 bar. After emulsification and homogenization three times, the liquid was placed in an 80°C water bath for 30 minutes to completely kill the living cells in the fermentation broth, thereby obtaining a Lactobacillus paracasei VHProbi E12 fermentation lysate.

[0044] Example 2 Inhibitory effect of Lactobacillus paracasei VHProbi E12 fermentation lysate on 5α-reductase activity

[0045] Buffer, 5α-reductase (10 mg / mL), Lactobacillus paracasei VHProbi E12 fermentation lysate, testosterone (4 μg / mL) and NADPH (10 mmol / L) were added to a 96-well plate in sequence. The sample volume is shown in Table 1. The total volume of the reaction system is 100 μL, the reaction temperature is 37 ° C, and the reaction time is 30 min. After the reaction is completed, 100 μL of methanol solution is added to 50 μL of the reaction solution to terminate the reaction. The supernatant is then centrifuged at 10,000 × g for 10 min and the test supernatant is taken. The testosterone content in the test sample is quantitatively determined by HPLC. The enzyme activity is measured by the consumption of the substrate testosterone. Each group is repeated 3 times.

[0046] Inhibition rate = [(BC-NC)-(BC-PC)] / (BC-NC) × 100%, where BC represents the testosterone content of the blank group, NC represents the testosterone content of the normal group, and PC represents the testosterone content of the reaction group to which the fermentation lysate of Lactobacillus paracasei VHProbi E12 was added.

[0047] Table 1 Reaction system liquid addition table

[0048]

[0049] The results showed that the Lactobacillus paracasei VHProbi E12 fermentation lysate had an inhibition rate of 51.29% ± 3.06% on 5α-reductase. This study has found a correlation between 5α-reductase and skin oil secretion. In sebaceous gland cells, androgens are converted into the more active dihydrotestosterone (DHT) through the combined action of 5α-reductase and NADPH. DHT further stimulates sebum secretion, thereby affecting the proliferation and differentiation of sebaceous gland cells. The Lactobacillus paracasei VHProbi E12 fermentation lysate was able to inhibit 5α-reductase activity, reduce skin lipid secretion, and alleviate oily skin.

[0050] Example 3 Inhibitory Effect of Lactobacillus paracasei VHProbi E12 Fermentation Lysate on Sebaceous Gland Cell Proliferation 3.1 Cell Culture

[0051] Sebomed medium supplemented with 5 ng / mL epidermal growth factor, 500 μg / mL streptomycin, 500 U / mL penicillin, and 10% fetal bovine serum was used to culture SZ95 human sebaceous gland cells in a 5% CO2 incubator at 37°C. Subsequent experiments were performed after the SZ95 human sebaceous gland cells reached 70% to 80% confluency. The final DMSO concentration was 0.2%.

[0052] 3.2 Inhibition test

[0053] SZ95 human sebaceous gland cells in good growth condition were cultured at a rate of 5×10 3 Cells were plated in 96-well plates and cultured in a 5% CO2 incubator at 37°C. The experiment was divided into a control group, a dihydrotestosterone (DHT) group, and a lysate group, with 12 replicates per group. After cell attachment, 10 μL of culture medium was added to the control group, 10 μM dihydrotestosterone was added to the DHT group, and 100 μL of dihydrotestosterone and 100 μL of Lactobacillus paracasei VHProbi E12 fermentation lysate were added to the lysate group. After the cells were cultured for 12 h, 24 h, and 36 h, 10 μL of CCK-8 solution was added to each well. After incubation for 2 h at 37°C, absorbance was measured at 450 nm using a microplate reader to measure cell viability.

[0054] The absorbance of cells in each group was Figure 1As shown, after 12 hours of incubation, the absorbances of the control, DHT, and lysate groups were 0.50, 0.67, and 0.57, respectively. Compared with the control group, the absorbance of the DHT group increased significantly (P < 0.01), indicating that DHT began to induce the proliferation of SZ95 human sebaceous gland cells. Compared with the DHT group, the absorbance of the lysate group decreased, but there was no significant difference between the two groups (P = 0.098), indicating that the Lactobacillus paracasei VHProbi E12 fermentation lysate can inhibit the DHT-induced proliferation of sebaceous gland cells. After 24 hours of incubation, the absorbances of the control, DHT, and lysate groups were 0.70, 1.00, and 0.91, respectively. Compared with the control group, the absorbance of the DHT group increased significantly (P < 0.01), indicating that DHT further enhanced the proliferation-inducing effect of SZ95 human sebaceous gland cells. Compared with the DHT group, the absorbance of the lysate group decreased, but there was no significant difference between the two groups (P = 0.186), indicating that the fermentation lysate of Lactobacillus paracasei VHProbi E12 can inhibit the proliferation of SZ95 human sebaceous gland cells induced by DHT. After 36 hours of incubation, the absorbances of the control group, DHT group, and lysate group were 0.98, 1.35, and 1.26, respectively. Compared with the control group, the absorbance of the DHT group increased, and the difference between the two groups was significant (P < 0.05). Compared with the DHT group, the absorbance of the lysate group decreased, and the difference between the two groups was significant (P < 0.01), indicating that the inhibitory effect of the fermentation lysate of Lactobacillus paracasei VHProbi E12 on the proliferation of SZ95 human sebaceous gland cells induced by DHT was further enhanced with prolonged incubation time. The above results show that Lactobacillus paracasei VHProbi E12 has an inhibitory effect on the proliferation of SZ95 human sebaceous gland cells, can reduce the excessive sebum secretion caused by excessive proliferation of sebaceous gland cells, and can inhibit the problem of excessive oil production of the skin.

[0055] Example 4 Inhibitory effect of Lactobacillus paracasei VHProbi E12 fermentation lysate on neutral lipid synthesis in SZ95 human sebaceous gland cells

[0056] 4.1 Cell culture

[0057] See Example 3 for cell culture.

[0058] 4.2 Neutral lipid synthesis inhibition test

[0059] SZ95 human sebaceous gland cells in good growth condition were cultured at 5×10 3Each well was inoculated with 100 μL of culture medium in a black, transparent 96-well plate and cultured at 37°C in a 5% CO2 incubator for 24 hours. The experiment was divided into a control group, a dihydrotestosterone (DHT) group, and a lysis solution group, with three replicates per group. After the cells adhered to the wall, 10 μL of culture medium was added to the control group, 10 μM dihydrotestosterone was added to the DHT group, and 100 μL of dihydrotestosterone and 100 μL of Lactobacillus paracasei VHProbi E12 fermentation lysate were added to the lysis solution group. Linoleic acid was added to all groups to induce SZ95 cell maturation at a final concentration of 100 μM. After 48 hours of cell culture, the supernatant was discarded and washed with PBS. Cells were stained with 15 μg / mL fluorescein diacetate (FDA) and 10 μg / mL Nile Red, respectively. After incubation at 37°C in the dark for 10 minutes, the cells were washed with PBS and the fluorescence intensities of FDA and Nile Red were measured using a microplate reader. The excitation and emission wavelengths for FDA were 494 nm and 523 nm, respectively, and for Nile Red were 485 nm and 565 nm, respectively. Lipid synthesis results were expressed as the ratio of Nile Red to FDA, and the percentage of the experimental group to the control group was used to reflect the relative lipid content in the cells.

[0060] The results are as follows Figure 2 As shown in the results, compared with the control group, the lipid synthesis rate in the dihydrotestosterone group was significantly increased (P<0.01), and the induction of linoleic acid significantly increased the lipid synthesis ability of SZ95 human sebocytes; compared with the dihydrotestosterone group, the lipid synthesis rate in the lysate group was significantly decreased (P<0.05), indicating that the fermentation lysate of Lactobacillus paracasei VHProbi E12 can significantly inhibit the synthesis of neutral lipids in SZ95 human sebocytes induced by linoleic acid, reduce the excessive secretion of sebum by sebaceous glands, and thus improve the problem of excessive oil production on the facial skin.

[0061] 4.3 Real-time fluorescence quantitative PCR detection

[0062] SZ95 cells in good growth condition were cultured at 6×10 4100 cells / well were inoculated in a 24-well plate and cultured at 37°C in a 5% CO2 incubator for 24 hours. The experiment was divided into a control group, a model group, a lysate group and a mixed group, with 3 replicates in each group. 0.1% DMSO was added to the control group, and 0.1mmol / L linoleic acid was used to induce SZ95 sebaceous gland cells to synthesize lipids in the model group and the mixed group. 250ul of Lactobacillus paracasei VHProbi E12 fermentation lysate was added to the lysate group and the mixed group. After incubation for 3 hours, the cells were collected and Trizol was added to extract total RNA for reverse transcription. The RNA was reverse transcribed into cDNA using a reverse transcription kit. After obtaining the template, GAPDH was used as the internal reference gene, and the PowerUp SYBR Green Master Mix kit and ABI 7500qPCR instrument were used to detect the expression levels of TRPV1 and AMPK mRNA. The relative expression of the target gene mRNA was expressed as 2 -ΔΔCt Reaction conditions: 95°C pre-denaturation for 30 s, 95°C denaturation for 5 s, 56°C annealing for 10 s, and 72°C extension for 25 s, for 40 cycles. Primer sequences are shown in Table 2.

[0063] Table 2 Primer sequences

[0064]

[0065] The results are as follows Figure 3 As shown in the results, compared with the control group, the expression levels of TRPV1 and AMPK mRNA in the model group were significantly decreased (P<0.01, P<0.05), and the expression levels of TRPV1 and AMPK mRNA in the lysate group were increased, but there was no statistical difference (P=0.19, P=0.12). Compared with the model group, the expression levels of TRPV1 and AMPK mRNA in the mixed group were increased, but there was no statistical difference (P=0.51, P=0.37).

[0066] The above results further confirmed that the fermentation lysate of Lactobacillus paracasei VHProbi E12 inhibits linoleic acid-induced excessive lipid synthesis in SZ95 human sebaceous gland cells by activating the TRPV1 and AMPK signaling pathways and increasing the expression levels of TRPV1 and AMPK mRNA, thereby effectively reducing the oil secretion of SZ95 human sebaceous gland cells and improving the problem of excessive oil production on the facial skin.

[0067] Example 5: Experiment on improving facial oiliness with Lactobacillus paracasei VHProbi E12

[0068] 5.1 Preparation of Lotion

[0069] A moisturizing lotion was prepared by combining 3% Lactobacillus paracasei VHProbi E12 fermentation lysate, 5% butylene glycol, 2% mineral oil, 1.5% capric glyceryl, 1.5% cetyl alcohol, 0.5% dimethicone, 0.5% peg-100 stearate, 0.4% 1,2-hexanediol, 0.3% hydroxyacetophenone, 0.25% phenoxyethanol, 0.2% glyceryl stearate, 0.15% carbomer, 0.13% arginine, 0.05% disodium edta and 79.52% water according to mass percentage for use by subjects.

[0070] 5.2 Detection

[0071] Fifty-six subjects aged 18-45 years with Fitzpatrick skin grades I-IV were recruited and asked to apply 0.6-0.8g of moisturizer twice daily, morning and evening, for 60 days. Before the test, the subjects sat quietly in an observation room for 20 minutes. On days 0 and 60, facial porphyrin levels were measured using the VISIA-CR instrument, and facial oil levels were measured using the CK-MAP instrument on days 0, 30, and 60. A questionnaire survey was also conducted.

[0072] 5.3 Results of improvement in subjects’ facial oiliness

[0073] The changes in the subjects' facial oil and porphyrin parameters are shown in Table 3. After the subjects used the moisturizer for 60 days, the problem of facial oiliness was significantly improved. The subjects' oil content was 61.00 on day 0, and decreased by 8.31% to 55.93 on day 30, with no significant difference compared with day 0 (P=0.07); on day 60, the subjects' oil content decreased by 15.31% to 51.67, with a significant difference compared with day 0 (P<0.01). The subjects' facial porphyrin content was 18.11 on day 0, and decreased by 32.85% to 12.16 on day 30, with a significant difference compared with day 0 (P=0.025); porphyrin is a metabolic product of some bacteria that parasitize in hair follicles and can reflect the secretion of sebaceous glands in the skin. The higher the porphyrin content in the skin test, the more vigorous the facial oil secretion, which is easy to block the hair follicles and promote the reproduction of acne bacteria. The changes in facial porphyrin content collected using the VISIA-CR instrument are shown as follows Figure 5 As shown, the subjects' facial porphyrin content was significantly reduced compared with the baseline period, and the oiliness of the skin was reduced.

[0074] The results of the study showed that after the subjects used the moisturizer containing Lactobacillus paracasei VHProbi E12 lysate for 60 days, the skin oil content, porphyrin content and melanin content were reduced, and the greasy condition of the skin was reduced.

[0075] Table 3 Changes in facial lipid and porphyrin parameters of subjects

[0076]

[0077] The results of the subject questionnaire survey are shown in Table 4. 80.4% of the subjects said that their skin oiliness was relieved, 94.6% of the subjects felt that their skin became smoother, 92.9% of the subjects felt that their skin was more delicate, 82.1% of the subjects had their skin itching relieved, 67.9% of the subjects had their skin pores improved, 83.9% of the subjects had their skin stability improved, 82.1% of the subjects had their facial complexion improved, and 98.2% of the subjects had their facial skin condition improved.

[0078] Table 4 Self-evaluation questionnaire on oil control efficacy

[0079]

[0080] In summary, the fermentation lysate of Lactobacillus paracasei VHProbi E12 inhibits 5α-reductase activity, reduces skin lipid secretion, and alleviates oily skin. It also significantly inhibits the proliferation of SZ95 human sebaceous gland cells, reducing excessive sebum secretion caused by excessive cell proliferation. Furthermore, the fermentation lysate of Lactobacillus paracasei VHProbi E12 activates the TRPV1 and AMPK signaling pathways, inhibiting linoleic acid-induced excessive lipid synthesis in SZ95 human sebaceous gland cells, thereby effectively reducing oil secretion in SZ95 human sebaceous gland cells and improving excessive facial oiliness. Human efficacy trials confirmed that after 60 days of using a moisturizer containing the fermentation lysate of Lactobacillus paracasei VHProbi E12, the oil content of the facial stratum corneum decreased by 15.30% and porphyrin decreased by 30.74%. 80.4% of the subjects reported a reduction in facial oiliness.

Claims

1. A strain of Lactobacillus paracasei ( Lacticaseibacillus paracasei ), characterized in that, The preservation number of the Lactobacillus paracasei is CCTCC NO: M2021587.

2. the application of the fermentation lysate of Lactobacillus paracasei described in claim 1 in improving the problem of excessive oil production of facial skin for non-disease treatment purpose.

3. The use according to claim 2, characterized in that The preparation method of the Lactobacillus paracasei fermentation lysate comprises the steps: (1) Fermentation broth preparation Under sterile conditions, Lactobacillus paracasei seed liquid was inoculated into a fermentation medium at a volume ratio of 3%, and cultured at 37° C. for 24 hours at a rotation speed of 100 rpm, a ventilation volume of 1 L / min, a tank pressure of 0.05 MPa, and a fermentation end point pH of 3.5-4.2 to prepare a Lactobacillus paracasei fermentation liquid; (2) Emulsification and homogenization of fermentation broth The Lactobacillus paracasei fermentation liquid was emulsified and homogenized using a homogenizer at a pressure of 1000 bar for 3 times, and then placed in an 80°C water bath for 30 minutes to completely kill the living cells in the fermentation liquid, thereby obtaining a Lactobacillus paracasei fermentation lysate.

4. The use according to claim 3, wherein the components and their contents in the fermentation medium are: sucrose 25 g / L, yeast powder 30 g / L, soy peptone 20 g / L, magnesium sulfate 0.10 g / L, and manganese sulfate 0.05 g / L.

5. Application of the fermentation lysate of Lactobacillus paracasei according to claim 1 in preparing a skin care product.

6. A skin care product, characterized in that: The skin care product comprises the fermentation lysate of the Lactobacillus paracasei according to claim 1.

7. The skin care product according to claim 6, wherein The skin care product further comprises any one or more of butylene glycol, mineral oil, glyceryl caprate, cetyl alcohol, dimethicone, peg-100 stearate, 1,2-hexanediol, hydroxyacetophenone, phenoxyethanol, glyceryl stearate, carbomer, arginine, and disodium edta.

8. The skin care product according to claim 7, wherein The components and their mass percentages in the skin care product are: 8% Lactobacillus paracasei VHProbi E12 fermentation lysate, 5% butylene glycol, 2% mineral oil, 1.5% capric glyceryl, 1.5% cetyl alcohol, 0.5% dimethicone, 0.5% PEG-100 stearate, 0.4% 1,2-hexanediol, 0.3% hydroxyacetophenone, 0.25% phenoxyethanol, 0.2% glyceryl stearate, 0.15% carbomer, 0.13% arginine, 0.05% disodium edta, and 79.52% water.

9. Use of the skin care product according to any one of claims 6 to 8 for improving excessive oil production on the facial skin for non-disease treatment purposes.

Images